1. Field of the Invention
The invention is directed to dynorphin oligopeptides with highly selective binding and biological activity at opioid receptors. More specifically, the invention is directed to cyclic and linear dynorphin A analogs which are highly potent and selective for kappa opioid receptors.
2. Discussion of the Background
Numerous oligopeptides have been identified in the central and peripheral nervous system. These oligopeptides display a variety of biological and pharmacological activities including analgesia, physiological dependence and tolerance, gut motility, etc. These nervous system oligopeptides include the highly active endorphins and enkephalns. Since the discovery of endogenous enkephalins there has been substantial research into the structure and activities of opioid oligopeptides and opioid non-peptides.
It is now well known that there are a multiplicity of opioid receptors in mammals, including the mu (.mu.), delta (.delta.), kappa (.kappa.) and possibly other subtypes of these receptors. These different receptors are thought to have different physiological roles and to interact differently with the various brain oligopeptides A continuing goal of research into opioid receptors and brain oligopeptides is the discovery of oligopeptide ligands which are highly specific for only one type of opioid receptor and which exhibit highly specific biological and pharmacological activity. Such peptides contribute substantially to the understanding of opioid receptors and have important biological and medical applications, including the management of pain without the undesirable side effects of morphine and other narcotics.
The enkephalins such as H--Tyr--Gly--Gly--Phe--Met--OH, i.e., Met.sup.5 -enkephalin, have modest selectivities for the .delta. receptor, while the dynorphins such as dynorphinl-.sub.1-11 (H--Tyr--Gly--Gly--Phe--Leu--Arg--Arg--Ile--Arg--Pro--Lys--OH) have modest specificity for the kappa receptor. The specificity of the various brain polypeptides for the individual receptors is thought to relate to the conformation and structure of the oligopeptides.
Shortly after the characterization of the endogenous enkephalins, attempts were made to determine their preferred conformations. By the use of various spectroscopic methods, such as X-ray crystallography and/or energy calculations, varying conclusions were reached. It is now accepted that the enkephalins are highly flexible molecules that can assume an ensemble of energetically preferred conformations.
There may be definite advantages for a biological system to utilize hormones or neurotransmitters having high conformational flexibility. Such advantages include the ability to utilize a variety of thermodynamically accessible pathways to ligand-receptor interactions, the ability of a specific hormone or neurotransmitter to assume different conformations, whereby the hormone or neurotransmitter could then effect different molecular pharmacological events, and the availability of different conformations for a specific ligand to permit binding to multiple types of receptors. For example, a single oligopeptide may variously bind to each of the mu, delta and kappa receptors with different specificity. Indeed, it is thought that the complexity of the pharmacological responses to the opioids may be due in part to their non-selective binding to opioid receptors.
In comparison to the mu and delta opioid receptors, research directed to kappa receptor oligopeptides has been limited, especially with peptide ligands. Structure-function studies have been primarily directed to the putative endogenous oligopeptides dynorphin A (Dyn A), dynorphin B (Dyn B) and .alpha.-neoendorphin. ##STR1##
Some structure-function relationships for dynorphin peptides have been determined (see for example, Chavkin, C.; Goldstein, A. (1981) Proc. Natl. Acad. Sci., USA 78, 6543-6547; Lemaire, S., Lafrance, L., Dumont, M. (1986), Int. J. Peptide Protein Res., 27, 300-305; Gairin, J.E., Gouarderes, C., Mazarguil, H., Alvinerie, P., Cros, J. (1984) Eur. J. Pharmacol., 106, 457-458; Gairin, J.E., Gout, R., Meunier, J.-C., and Cros, J. (1988), J. Pharmacol. Exp. Ther., 245, 995-1001).
Thus far, the most potent and selective kappa opioid receptor agonists have been several non-peptide analogs of N-methyl-N-[(pyrrolidinyl)-cyclohexyl]benzeneacetamide. FIG. 1 shows the structures of U-50488 and related analogs. The analog U-50488 is a racemic mixture while U-69593 is enantiomerically pure. U-50488 and its analogs do not display morphine-type physical dependence, urogenic activity or respiratory depression and appear to prevent the development of tolerance to morphine analgesia. However, use of these compounds is disadvantageous in that they induce tolerance, cause water diuresis and have been implicated in the production of psychotomimesis.
The most potent and selective peptidic opioid receptor agonist which has been reported is [D-pro.sup.10 ]Dyn A.sub.1-11 -OH (DPDYN). See Gairin et al., Eur. J. Pharmacol, loc. cit. DPDYN exhibits high .kappa. vs. .mu. and .kappa. vs .delta. selectivities with a reported binding affinity of about 0.032 nM against the kappa ligand [.sup.3 H]Bremazocine. When administered i.c.v., DPDYN did not show any activity against thermal stimulus but, in contrast, produced a dose-related effect against chemical pain (Gairin et al., J. Pharmacol. Exp. Ther. loc. cit.).
Attempts have been made to investigate the secondary structure of Dyn-A using various spectroscopic methods. Infrared attenuated total reflection spectroscopy and capacitance minimization have been used to study the secondary structure, orientation and accumulation of DYN A.sub.1-13 molecules on the surface of planar phosphocholine derived membranes (D. Erne, et al. (1985) Biochemistry, 24:4161-4263). Erne et al. proposed that the peptide assumes a helical structure extending from Tyr.sup.1 to Pro.sup.10 oriented perpendicularly to the membrane surface, while the remaining C-terminal residues adopt a random coil conformation.
To study the aqueous phase secondary structure of Dyn A.sub.1-13, FT-infrared spectroscopic studies in H.sub.2 O and D.sub.2 were utilized in conjunction with proton NMR deuterium exchange studies (V. Renugopalakrishnan et al., (1988), Biochem. Biophys. Res. Commun., 151:1220-1225). The peptide NH groups appeared to be solvent accessible which was suggestive of an essentially extended structure with a periodically interwoven unordered structure. These results are consistent with previous Raman spectroscopic experiments (Rapaka, R.S., Renugopalakrishnan,. V., Collete, T.W., Dobbs, J.C., Carreira, L.A., and Bhatnagar, R.S. (1987), Int. J. Peptide Protein Res., 30, 284-287; 2D-NOESY spectra (Renugopalakrishnan, V., Rapaka, R.S., Huang, S.-G., Moore, S. and Hutson, T.B. (1988), Biochem. Biophys. Res. Commun., 151, 1220-1225) and extensive lD and 2D NMR studies (Zhou, N. and Gibbons, W.A. (1986), J. Chem. Soc. Perkin Trans. II, 637-644). Additional spectroscopic experiments supporting the existence of an extended and/or random conformation of Dyn A in an aqueous environment include fluorescence energy transfer studies (Schiller, P.W., (1983), Int. J. Peptide Protein Res., 21, 307-312) and circular dichroism studies (Kojro, E., Gwizdala, E., Grzonka, Z. (1987), Polish J. Chemistry, 61, 415-424).
Additionally, the secondary structure of dynorphin oligopeptides has been studied through the use of conformationally constrained analogs. The first conformationally constrained dynorphin A analog reported was the cyclic disulfide ##STR2## (Schiller, P.W., Eggiman, B., and Nguyen, T.M.-D, (1982), Life Sci., 31, 1777-1780). This compound exhibits high biological activity and a high affinity for the mu receptor. Cyclic lactams of Dyn A such as ##STR3## have also been prepared (Schiller, P.W., Nguyen, T.M.-D., Lemieux, C. (1988), Tetrahedron, 44, 733-743). These cyclic lactams show varying degrees of biological activity but generally low or negligable interaction with the kappa receptor. It has generally been concluded that these cyclic lactams of Dyn A do not have kappa selectivity.
A continuing need exists for new oligopeptides with improved receptor selectivity enhanced biological activity, and selectivity for the brain or peripheral kappa receptor sites.